Effort will be concentrated upon cultivation in defined media of functional cells from mouse tissues, including liver, kidney, and prostate. The approaches will be (1) by cell secretion, either in primary or later stages of culture, to reduce or eliminate non-epithelial cells. Methods for this will include differential cell attachment to a substrate, a property that is dependent both upon the type of cell and upon the composition and treatment of the attachment surface: use of inhibitors of firbroblasts (e.g. hydrocortisone, cis-4-hydroxy-L-proline, Gentamicin) or of fibroblasts and lymphocytes (e.g. D-valine), or target-specific hormones that are promoters of particular types of epithelial cell. We shall (2) continue refinement of the environment of the cell culture system, including adjustments in medium composition, with special attention to the substitution for sodium bicarbonate of non-bicarbonate buffers, either sodium glycerophosphate or, where that is not appropriate, other. We shall examine, in media that are buffered other than with bicarbonate, the needs of the cells for a source of CO2, either provided as such or generated endogenously, e.g. from pyruvate. Substitutes for L-glutamine, to enable the media to be fully autoclavable, will be tested. Other aspects of the environment to be examined are ion balance, osmolality (especially for kidney cells), where several cell types may be selected at different osmotic levels corresponding to the osmotic pressures to which they are exposed in vivo. Functions (3) appropriate to the cell types studied will be tested, e.g. serum protein production by liver cells, and enzymes specific to each cell type.